Cloned (Comment) | Organism |
---|---|
gene RSP_3671, sequence comparisons and phylogenetic tree, recombinant expression in Escherichia coli strain BL21(DE3) | Cereibacter sphaeroides |
Protein Variants | Comment | Organism |
---|---|---|
R118W | site-directed mutagenesis, the unique hydrogen bond between Arg118 and O4 of D-fructose is broken when Arg118 is mutated to Trp, the mutation improves the substrate recognition and activity of the enzyme. The mutant enzyme RsDTE_R118W shows decreased catalytic activity compared to the wild-type enzyme toward D-fructose, the kcat/Km for D-tagatose is about twofold higher than for D-psicose. Mutant R118W shows 1.5fold higher catalytic efficiency toward D-tagatose than the wild-type | Cereibacter sphaeroides |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | - |
Cereibacter sphaeroides | |
Cu2+ | high inhibition at 0.1 mM | Cereibacter sphaeroides | |
EDTA | - |
Cereibacter sphaeroides | |
Fe2+ | high inhibition at 0.1 mM | Cereibacter sphaeroides | |
Fe3+ | - |
Cereibacter sphaeroides | |
Ni2+ | almost complete inhibition at 0.1 mM | Cereibacter sphaeroides | |
Zn2+ | - |
Cereibacter sphaeroides |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Cereibacter sphaeroides | |
0.98 | - |
D-tagatose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
78 | - |
D-fructose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
137.9 | - |
D-tagatose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
148.1 | - |
D-sorbose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
162.1 | - |
D-fructose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
192.9 | - |
D-psicose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
215.2 | - |
D-psicose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
359.4 | - |
D-sorbose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Co2+ | dependent on, activates at 0.1 mM, metalloenzyme | Cereibacter sphaeroides | |
Mn2+ | dependent on, activates at 0.1 mM, metalloenzyme, Mn2+ is the most effective activating divalent cation | Cereibacter sphaeroides | |
additional information | no effect on enzyme activity by Mg2+ at .1 mM | Cereibacter sphaeroides |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-fructose | Cereibacter sphaeroides | - |
D-psicose | - |
r | |
D-fructose | Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | - |
D-psicose | - |
r | |
D-tagatose | Cereibacter sphaeroides | - |
D-sorbose | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Cereibacter sphaeroides | Q3IW04 | - |
- |
Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | Q3IW04 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
D-fructose | - |
Cereibacter sphaeroides | D-psicose | - |
r | |
D-fructose | best substrate for the recombinant wild-type enzyme | Cereibacter sphaeroides | D-psicose | - |
r | |
D-fructose | - |
Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | D-psicose | - |
r | |
D-fructose | best substrate for the recombinant wild-type enzyme | Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | D-psicose | - |
r | |
D-psicose | - |
Cereibacter sphaeroides | D-fructose | - |
r | |
D-psicose | - |
Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | D-fructose | - |
r | |
D-sorbose | - |
Cereibacter sphaeroides | D-tagatose | - |
r | |
D-sorbose | - |
Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | D-tagatose | - |
r | |
D-tagatose | - |
Cereibacter sphaeroides | D-sorbose | - |
r | |
D-tagatose | best substrate for the recombinant enzyme mutant R118W | Cereibacter sphaeroides | D-sorbose | - |
r | |
additional information | RsDTE wild-type shows lower Michaelis-Menten constant (Km), lower turnover number (kcat), but higher catalytic efficiency (kcat/Km) values for D-fructose than for D-psicose. The kcat/Km for D-fructose is 5.5fold higher than for D-psicose, indicating that enzyme RsDTE highly catalyzes D-fructose, although it is a D-tagatose 3-epimerase | Cereibacter sphaeroides | ? | - |
? | |
additional information | RsDTE wild-type shows lower Michaelis-Menten constant (Km), lower turnover number (kcat), but higher catalytic efficiency (kcat/Km) values for D-fructose than for D-psicose. The kcat/Km for D-fructose is 5.5fold higher than for D-psicose, indicating that enzyme RsDTE highly catalyzes D-fructose, although it is a D-tagatose 3-epimerase | Cereibacter sphaeroides ATCC 17023 / 2.4.1 / NCIB 8253 / DSM 158 | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
RsDTE | - |
Cereibacter sphaeroides |
RSP_3671 | - |
Cereibacter sphaeroides |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
35 | - |
- |
Cereibacter sphaeroides |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
20 | 70 | half-maximal activity at 20°C, near inactivation at 70°C | Cereibacter sphaeroides |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
8.3 | - |
D-sorbose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
15.36 | - |
D-sorbose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
20.47 | - |
D-psicose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
26.04 | - |
D-psicose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
41.05 | - |
D-fructose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
42.6 | - |
D-fructose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
48.31 | - |
D-tagatose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
51.25 | - |
D-tagatose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8.5 | - |
- |
Cereibacter sphaeroides |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
5.5 | 11 | activity range | Cereibacter sphaeroides |
General Information | Comment | Organism |
---|---|---|
evolution | the enzyme belongs to the D-tagatose 3-epimerase (D-TE) family | Cereibacter sphaeroides |
additional information | the O1 of substrate D-fructose forms hydrogen bonds with His192 and Glu162 to help the correct metal coordination of the substrate. The O2 forms hydrogen bonds with His192 and Arg221. Glu156 forms hydrogen bonds with O3, and Glu250 directs its OE2 atom to a hydrogen atom attached to C3. Because D-fructose has the same configurations of C1, C2 and C3 as D-tagatose, the interactions between D-fructose at the 1-, 2- and 3-positions and the enzyme are very similar to those in other DTE/DPE family enzymes. Residue R118 forms a hydrogen bond with O4 of D-fructose and may regulate the substrate specificity. The strengthened hydrophobic interaction may attribute to the recognition of D-tagatose, D-psicose, and D-sorbose. Enzyme homology modeling and structure comparisons, overview | Cereibacter sphaeroides |
physiological function | the purified D-tagatose 3-epimerase from Rhodobacter sphaeroides catalyzes the epimerization of D-fructose to D-psicose at the C3 position | Cereibacter sphaeroides |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.02 | - |
D-sorbose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.1 | - |
D-sorbose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.1 | - |
D-psicose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.14 | - |
D-psicose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.25 | - |
D-fructose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.35 | - |
D-tagatose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.52 | - |
D-tagatose | recombinant mutant R118W, pH 8.5, 35°C | Cereibacter sphaeroides | |
0.55 | - |
D-fructose | recombinant wild-type enzyme, pH 8.5, 35°C | Cereibacter sphaeroides |